Despite its availability, affordability and efficacy, there is a concern that depo medroxyprogesterone acetate (depo-MPA, DMPA) increases the susceptibility of women to HIV-1. Accordingly, we seek an improved knowledge of the effects of contraceptive progestins on HIV-1 transmission. Contraceptive steroid effects are highly dependent on receptor-binding and local concentration and macaque studies illustrate the importance of the endocervix in SIV transmission. Still, there are no data on local endocervical concentrations of any contraceptive progestins in women using these medications. Systemically-delivered DMPA is predicted to produce low genital tract concentrations of MPA. MPA's strong affinity for the glucocorticoid receptor (GR) may enable it to modulate immune and physical epithelial barrier functions that promote HIV transmission in the endocervix. As alternatives to DMPA, levonorgestrel (LNG), nomegestrel acetate (NOMAC) and the PR modulator, ulipristal (UL), have poor affinity for the GR and may be delivered in high local concentrations to the genital tract. We hypothesize that high levels of LNG, NOMAC or UL, unlike low levels of MPA, protect the physical and innate epithelial barrier of the endocervix and decrease HIV-1 transmission at this site. We will measure progestin levels in the endocervix of women using DMPA, an LNG-releasing IUD and a progestin-containing vaginal ring and assess effects of progestin type and delivery method on the physical and immune barrier of cervical mucus to HIV-1. Next, we will expose polarized, hormone-responsive endocervical epithelial cells to physiologic levels of contraceptive progestins and follow changes in monolayer integrity, the innate immune response and HIV-1 transmission across this barrier. Lastly we will investigate the susceptibility of endocervical T cells in women using DMPA and the LNG-IUD to HIV-1 and test the hypothesis that epithelial mediators increase T cell susceptibility to HIV-1 under low levels of MPA but not high levels of LNG, NOMAC or UL. Understanding the mechanisms underlying increased HIV-1 transmission in women using DMPA and the differential effects of alternative progestins and delivery methods on transmission events will allow informed recommendation of the safest contraceptive methods for use in at-risk women.